Prodrugs for antimicrobial amidines

ABSTRACT

A methods of treating an infection comprises administering a therapeutically effective amount of a compound described by the Formula (I)

This application is a divisional of U.S. patent application Ser. No.09/918,787 filed Jul. 31, 2001, now abandoned which is a divisional ofU.S. patent application Ser. No. 09/612,138 filed Jul. 7, 2000, now U.S.Pat. No. 6,486,200 herein incorporated by reference in their entirety,which is based on and claims priority to U.S. Provisional PatentApplication Serial No. 60/142,826 filed Jul. 8, 1999, hereinincorporated by reference in its entirety.

GRANT STATEMENT

The present invention was made with Government support under GrantNumber 5-U19-A133363 from the National Institutes of Health. The U.S.Government has certain rights to this invention.

FIELD OF THE INVENTION

The invention generally relates to methods for treating infections.

BACKGROUND OF THE INVENTION

A microbial infection such as, for example, Pneumocystis cariniipneumonia (PCP), is believed to be one of the leading causes of death inpatients suffering from AIDS. Pentamidine [i.e.,1,5-bis(4-amidinophenoxy)pentane] has been used as a therapeutic agentfor the treatment of PCP by intravenous infusion and as a prophylacticagent by aerosol dosage. However, the use of this drug may bepotentially disadvantageous in that it might be toxic and contribute tohypotension, hypoglycemia, and cardiac arrhythmias experienced by thepatient taking pentamidine.

Recent efforts have focused on developing other compounds forpotentially treating PCP. A number of aromatic diamidines have displayedpotential anti-PCP activity as reported in Boykin D. W., et al., J. Med.Chem., 1995, pp. 912-916; Tidwell, R. R. et al., Antimicrob. AgentsChemother. 1993, 37, p. 1713; Lombardy, R. L. et al., J. Med. Chem.1996, 39, p. 1452; and Kumar, A. et al., J. Med. Chem., 1996, 31, p.767. Notwithstanding any advantages that these drugs may possess, theymay be potentially undesirable since the drugs often exhibit low oralbioavailability.

Chemical modification of drugs into prodrugs can potentially improvephysiochemical properties such as water solubility, lipophilicity,transport of drug to the site of action, and presystemic degradation,thus improving oral bioavailability. See Bundgaard, H., In Design ofProdrugs; Bundgaard, H.;Ed.; Elsevier: Amsterdam, The Netherlands, 1985;pp. 1-92; and Bundgaard, H., In A Textbook of Drug Design andDevelopment, Krogsgaard-Larsen, P.; Bundgaard, H.; Ed.; Harwood AcademicPubl. Switzerland, 1991, pp. 113-191. A number of reports exist on theprodrug modification of carboxyl, hydroxyl, thiols, and amino compounds.See, for example, Friis, G. J., et al., In A Textbook of Drug Design andDevelopment, 2^(nd) Ed., Krogsgaard-Larsen, P., Liljefors, T. Madsen,U.; Ed.; Overseas Pub: Amsterdam, The Netherlands, 1996, pp. 351-385;Digenis, G. A., et al., Drug latentiation, Handbook of ExperimentalPharmacology, 1975, 28, pp. 86-112. Moreover, Weller et al. (J. Med.Chem., 1996, 39, pp. 3139-3146) propose employing amidoximes andcarbamate derivatives of mono-amidines as prodrugs in order topotentially provide improved oral availability for fibrogen antagonists.In addition, Boykin, D. W., et al., (Bioorg. Med. Chem. Lett., 1996, 6,pp. 3017-3020) have reported that bis-amidoxime and O-methylamidoximemay be effective anti-PCP agents on both oral and intravenousadministration. U.S. Pat. No. 5,723,495 to Hall et al. proposesadministering a bis-benzamidoxime to a patient for treating Pneumocystiscarinii.

Notwithstanding the above efforts, there remains a need in the art toprovide drugs that display improved activity.

SUMMARY OF THE INVENTION

A method of combating an infection to a subject in need of such atreatment is disclosed. The method comprises administering to thesubject a compound of the formula (I):

wherein:

X may be O, S, or NR′ wherein R′ is H or loweralkyl;

R₁ and R₂ may be independently selected from the group consisting of H,loweralkyl, oxyalkyl, alkoxyalkyl, cycloalkyl, aryl, hydroxyalkyl,aminoalkyl, and alkylaminoalkyl;

R₃ and R₄ are each independently selected from the group consisting ofH, loweralkyl, halogen, oxyalkyl, oxyaryl, and oxyarylalkyl;

R₅ is represented by a formula selected from the group consisting of:

wherein:

X₁, X₂, and X₃ are independently selected from O and S; and

R₆ and R₇ are independently selected from the group consisting ofloweralkyl, aryl, alkylaryl, oxyaryl, an ester-containing substituent,and oxyalkyl;

or a pharmaceutically acceptable salt thereof, and wherein said compoundof Formula (I) is administered in an amount to treat the infection.

Preferably, R₆ and R₇ are independently selected from the groupconsisting of:

CH₃, CH₂CCl₃, CH₂CH₃,

In one preferred embodiment, each of the substituents present on thecompound of formula (I) represented by the formula:

are present on the para positions of the aromatic groups on formula (I),although these substituents may be present in the meta positions.

The invention also discloses pharmaceutical compounds represented by theformula (I) described herein and pharmaceutically acceptable saltsthereof, as well as pharmaceutical formulations comprising thepharmaceutical compounds of formula (I) and pharmaceutically acceptablecarriers.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 illustrates a reaction in the synthesis of carbamates from2,5-bis(4-amidinophenyl)furan.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The present invention now will be described more fully hereinafter withreference to the accompanying specification and examples, in whichpreferred embodiments of the invention are shown. This invention may,however, be embodied in many different forms and should not be construedas limited to the embodiments set forth herein. Rather, theseembodiments are provided so that this disclosure will be thorough andcomplete, and will fully convey the scope of the invention to thoseskilled in the art.

As used herein, the term “lower alkyl” refers to C1 to C4 linear orbranched alkyl, such as methyl, ethyl, propyl, butyl, isopropyl,sec-butyl, and tert-butyl. The term “halogen” has its conventionalmeaning and refers to fluorine, chlorine, bromine, and iodine. The term“cycloalkyl” as used herein refers to C3 to C6 cyclic alkyl, such ascyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. The term “aryl” asused herein refers to C3 to C10 cyclic aromatic groups such as phenyl,naphthyl, and the like, and includes substituted aryl groups such as,but not limited to, tolyl. The term “hydroxyalkyl” as used herein refersto C1 to C4 linear or branched hydroxy-substituted alkyl, i.e., —CH₂OH,—(CH₂)₂OH, etc. The term “aminoalkyl” as used herein refers to C1 to C4linear or branched amino-substituted alkyl, wherein the term “amino”refers to the group NR′R″, wherein R′ and R″ are independently selectedfrom H or lower alkyl as defined above, i.e., —NH₂, —NHCH₃, —N(CH₃)₂,etc. The term “oxyalkyl” as used herein refers to C1 to C4oxygen-substituted alkyl, i.e., —OCH₃, and the term “oxyaryl” as usedherein refers to C3 to C10 oxygen-substituted cyclic aromatic groups.The term “alkoxyalkyl” as used herein refers to C1 to C4 linear orbranched alkoxy, such as methoxy, ethoxy, propyloxy, butyloxy,isopropyloxy, and t-butyloxy. The term “ester-containing substituent”refers to a substituent that may be directed linked to the compound offormula (I) via the single bond that is present directly off of theoxygen atom contained in the ester group or may be of the formula:

wherein R′ and R″ may be the same or different and can be substituted orunsubstituted alkyl that may be saturated or unsaturated. It should beappreciated that the various groups referred to above may be substitutedor unsubstituted with various functional groups known to one skilled inthe art.

As noted above, the methods of the present invention are useful fortreating microbial infections such as P. carinii and Giardia lamblia.The compounds may also be useful in treating fungal infections such asCandida albicans, Cryptococcus neoformans, Aspergillus fumigatus,Fusarium solani, and combinations thereof. The methods of the inventionare useful for treating these conditions in that they inhibit the onset,growth, or spread of the condition, cause regression of the condition,cure the condition, or otherwise improve the general well-being of asubject afflicted with, or at risk of contracting the condition.

Subjects to be treated by the methods of the present invention aretypically human subjects, although the methods of the present inventionmay be useful with any suitable subject known to those skilled in theart.

As noted above, the present invention provides pharmaceuticalformulations comprising the aforementioned active compounds, orpharmaceutically acceptable salts thereof, in pharmaceuticallyacceptable carriers for oral, intravenous, or aerosol administration asdiscussed in greater detail below. Also, the present invention providessuch compounds or salts thereof which have been lyophilized and whichmay be reconstituted to form pharmaceutically acceptable formulationsfor administration, as by intravenous or intramuscular injection.

The therapeutically effective dosage of any specific compound, the useof which is in the scope of present invention, will vary somewhat fromcompound to compound, and patient to patient, and will depend upon thecondition of the patient and the route of delivery. As a generalproposition, a dosage from about 0.1 to about 50 mg/kg will havetherapeutic efficacy, with all weights being calculated based upon theweight of the active compound, including the cases where a salt isemployed. Toxicity concerns at the higher level may restrict intravenousdosages to a lower level such as up to about 10 mg/kg, with all weightsbeing calculated based upon the weight of the active base, including thecases where a salt is employed. A dosage from about 10 mg/kg to about 50mg/kg may be employed for oral administration. Typically, a dosage fromabout 0.5 mg/kg to 5 mg/kg may be employed for intramuscular injection.Preferred dosages are 1 μmol/kg to 50 μmol/kg, and more preferably 22μmol/kg and 33 μmol/kg of the compound for intravenous or oraladministration. The duration of the treatment is usually once per dayfor a period of two to three weeks or until the condition is essentiallycontrolled. Lower doses given less frequently can be usedprophylactically to prevent or reduce the incidence of recurrence of theinfection.

In accordance with the present method, pharmaceutically active compoundsas described herein, or pharmaceutically acceptable salts thereof, maybe administered orally as a solid or as a liquid, or may be administeredintramuscularly or intravenously as a solution, suspension, or emulsion.Alternatively, the compounds or salts may also be administered byinhalation, intravenously or intramuscularly as a liposomal suspension.When administered through inhalation the active compound or salt shouldbe in the form of a plurality of solid particles or droplets having aparticle size from about 0.5 to about 5 microns, and preferably fromabout 1 to about 2 microns.

The present invention also provides a pharmaceutical compositionsuitable for intravenous or intramuscular injection. The pharmaceuticalcomposition comprises a compound of formula (I) described herein, or apharmaceutically acceptable salt thereof, in any pharmaceuticallyacceptable carrier. If a solution is desired, water is the carrier ofchoice with respect to water-soluble compounds or salts. With respect tothe water-insoluble compounds or salts, an organic vehicle, such asglycerol, propylene glycol, polyethylene glycol, or mixtures thereof,may be suitable. In the latter instance, the organic vehicle may containa substantial amount of water. The solution in either instance may thenbe sterilized in a suitable manner known to those in the art, andtypically by filtration through a 0.22 micron filter. Subsequent tosterilization, the solution may be dispensed into appropriatereceptacles, such as depyrogenated glass vials. Of course, thedispensing is preferably be done by an aseptic method. Sterilizedclosures may then be placed on the vials and, if desired, the vialcontents may be lyophilized.

In addition to compounds of formula (I) or their salts, thepharmaceutical compositions may contain other additives, such aspH-adjusting additives. In particular, useful pH-adjusting agentsinclude acids, such as hydrochloric acid, bases or buffers, such assodium lactate, sodium acetate, sodium phosphate, sodium citrate, sodiumborate, or sodium gluconate. Further, the compositions may containmicrobial preservatives. Useful microbial preservatives includemethylparaben, propylparaben, and benzyl alcohol. The microbialpreservative is typically employed when the formulation is placed in avial designed for multidose use. Of course, as indicated, thepharmaceutical compositions of the present invention may be lyophilizedusing techniques well known in the art.

In yet another aspect of the present invention, there is provided aninjectable, stable, sterile composition comprising a compound of Formula(I), or a salt thereof, in a unit dosage form in a sealed container. Thecompound or salt is provided in the form of a lyophilizate which iscapable of being reconstituted with a suitable pharmaceuticallyacceptable carrier to form a liquid composition suitable for injectionthereof into a subject. The unit dosage form typically comprises fromabout 10 mg to about 10 grams of the compound or salt, When the compoundor salt is substantially water-insoluble, a sufficient amount ofemulsifying agent which is physiologically acceptable may be employed insufficient quantity to emulsify the compound or salt in an aqueouscarrier. One such useful emulsifying agent is phosphatidyl choline.

Other pharmaceutical compositions may be prepared from thewater-insoluble compounds disclosed herein, or salts thereof, such asaqueous base emulsions. In such an instance, the composition willcontain a sufficient amount of pharmaceutically acceptable emulsifyingagent to emulsify the desired amount of the compound or salt thereof.Particularly useful emulsifying agents include phosphatidyl cholines,and lecithin.

Further, the present invention provides liposomal formulations of thecompounds disclosed herein and salts thereof. The technology for formingliposomal suspensions is well known in the art. When the compound orsalt thereof is an aqueous-soluble salt, using conventional liposometechnology, the same may be incorporated into lipid vesicles. In such aninstance, due to the water solubility of the compound or salt, thecompound or salt will be substantially entrained within the hydrophiliccenter or core of the liposomes. The lipid layer employed may be of anyconventional composition and may either contain cholesterol or may becholesterol-free. When the compound or salt of interest iswater-insoluble, again employing conventional liposome formationtechnology, the salt may be substantially entrained within thehydrophobic lipid bilayer which forms the structure of the liposome. Ineither instance, the liposomes which are produced may be reduced insize, as through the use of standard sonication and homogenizationtechniques.

Of course, the liposomal formulations containing the compounds disclosedherein or salts thereof, may be lyophilized to produce a lyophilizatewhich may be reconstituted with a pharmaceutically acceptable carrier,such as water, to regenerate a liposomal suspension.

Pharmaceutical formulations are also provided which are suitable foradministration as an aerosol, by inhalation. These formulations comprisea solution or suspension of a desired compound described herein or asalt thereof, or a plurality of solid particles of the compound or salt.The desired formulation may be placed in a small chamber and nebulized.Nebulization may be accomplished by compressed air or by ultrasonicenergy to form a plurality of liquid droplets or solid particlescomprising the compounds or salts. The liquid droplets or solidparticles should have a particle size in the range of about 0.5 to about10 microns, more preferably from about 0.5 to about 5 microns. The solidparticles can be obtained by processing the solid compound or a saltthereof, in any appropriate manner known in the art, such as bymicronization. Most preferably, the size of the solid particles ordroplets will be from about 1 to about 2 microns. In this respect,commercial nebulizers are available to achieve this purpose. Thecompounds may be administered via an aerosol suspension of respirableparticles in a manner set forth in U.S. Pat. No. 5,628,984, thedisclosure of which is incorporated herein by reference in its entirety.

Preferably, when the pharmaceutical formulation suitable foradministration as an aerosol is in the form of a liquid, the formulationwill comprise a water-soluble compound or a salt thereof, in a carrierwhich comprises water. A surfactant may be present which lowers thesurface tension of the formulation sufficiently to result in theformation of droplets within the desired size range when subjected tonebulization.

As indicated, the present invention provides both water-soluble andwater-insoluble compounds and salts thereof. As used in the presentspecification, the term “water-soluble” is meant to define anycomposition which is soluble in water in an amount of about 50 mg/mL, orgreater. Also, as used in the present specification, the term“water-insoluble” is meant to define any composition which hassolubility in water of less than, about 20 mg/mL. For certainapplications, water soluble compounds or salts may be desirable whereasfor other applications water-insoluble compounds or salts likewise maybe desirable.

In another aspect, the invention relates to a process for making apharmaceutically active bis-aryl carbamate represented by formula (I).The process comprises reacting an aryl carbonate with bis-amidine in thepresence of an organic solvent to form the bis-aryl carbamate.

In one embodiment, the aryl carbonate may be represented by the formula:

wherein:

R is represented by:

wherein X is selected from the group consisting of H, NO₂, F, and OCH₃;and wherein R′ is selected from the group consisting of CH₃, CH₃CH₂,CH₂CCl₃, CH(OAc)CH₂, CH₂C₆H₅, and

wherein X is selected from the group consisting of H, NO₂, F, and OCH₃.The aryl carbonate may be a symmetrical aryl carbonate. Specificexamples of aryl carbonates include, but are not limited to, diphenylcarbonate, bis(4-fluorophenyl)carbonate, bis(4-methoxyphenyl)carbonate,benzyl4-nitrophenylcarbonate, 4-nitrophenyl thioethyl carbonate, and4-nitrophenyl-2,2,2-trichloroethyl carbonate, methyl 4-nitrophenylcarbonate, bis (3-fluorophenyl)carbonate, ethyl 4-nitrophenyl carbonate,(4-methyl-2-oxo-1,3-dioxol-4-en-5-yl)methyl 4-nitrophenyl carbonate, and1-acetoxyethyl 4-nitrophenyl carbonate.

Examples of the pharmaceutically active bis-aryl carbamate that may beformed by the process of the invention include, but are not limited to,2,5-bis[4-(N-2,2,2-trichloroethoxycarbonyl)amidinophenyl]furan,2,5-bis[4-(N-thioethylcarbonyl)amidinophenyl]furan,2,5-bis[4-(N-benzyloxy-carbonyl)amidinophenyl]furan,2,5-bis[4-(N-phenoxycarbonyl)amidino-phenyl]furan,2,5-bis[4-(N-(4-fluoro)phenoxycarbonyl)amidinophenyl]furan,2,5-bis[4-(N-(4-methoxy)phenoxycarbonyl)amidinophenyl]furan,2,5-bis[4(1-acetoxyethoxycarbonyl)amidinophenyl]furan, and 2,5-bis[4-(N-(3-fluoro)phenoxycarbonyl)amidinophenyl] furan.

Preferred organic solvents that may be employed in the process of theinvention include, but are not limited to, dimethyl formamide,tetrahydrofuran/CH₃CN, and dioxane. Typically, tetrahydrofuran/CH₃CN isemployed in the presence of a base such as, but not limited to,diisopropylethylamine and triethylamine.

In addition to the above, the compounds described herein may be formedby various methods such as, for example, those described in Weller, T.,et al., J. Med. Chem. 1996, 39, 3139-3146. Such a method typicallyrelates to preparing a carbamate from amines of amidines involving thereaction of the base with an appropriate chloroformate in the presenceof a base, typically aqueous sodium hydroxide or sodium/potassiumbicarbonate. Nonetheless, since this method may suffer from potentialdrawbacks, an alternative method is disclosed herein for preparingcarbamates from amidines by reaction with carbonates, and in particulararyl-alkyl and aryl—aryl carbonates. Traditionally carbonates have beenprepared by the reaction of tertiary amines and chloroformates. SeeOlofson, R. A., et al., J. Org. Chem. 1984, 49, pp. 2081-2082; andOlofson, R. A. Pure and Appl. Chem. 1988, 60, pp. 1715-1724.Additionally, the dealkylation of tertiary aliphatic amines with phenylchlorothionoformate has been reported in Millan D. S., et al.,Tetrahedron Lett. 1998, 39, pp. 4387-4390.

A method for preparing carbonates is disclosed herein in which pyridineis used as a base. As an example of the method, the 4-nitrophenylalkylcarbonates described herein were prepared by reacting 4-nitrophenol withthe corresponding alkyl or arylchloroformates in methylene chloride(CH₂Cl₂) using pyridine as a base. (1-acetyloxy)ethyl-4-nitrophenylcarbonate was prepared from 1-chloroethyl-4-nitrophenyl carbonateaccording to published procedures. See Alexander, J. et al., J. Med.Chem., 1991, 34, pp. 78-81; and Lin, Y. I., et al., Bioorg. Med. Chem.Lett., 1997, 7, pp. 1811-1816. Symmetrical carbonates (i.e., diphenyland bis(4-fluoro)- and bis(4-methoxy)phenylcarbonates described as20-22) were synthesized by reacting phenol, 4-fluorophenol, and4-methoxyphenol with phenyl, 4-fluorophenyl, and 4-methoxypehnylchloroformates respectively in a pyridine/CH₂Cl₂ medium. 4-nitrophenyl(5-methyl-2-oxo-1,3-dioxo-4-ene-1-yl) methyl carbonate (23) wassynthesized from commercially available 4,5-dimethyl-1,3-dioxol-2-one bya four-step process as outlined in Sakamoto, F. et al., Chem. Pharm.Bull., 1984, 32, pp. 2241-2248. A modification to this procedure hasbeen proposed in the bromination step which ultimately leads to thecarbonate 23. The bromination of dimethyl dioxolone withN-bromosuccinimide in carbon tetrachloride in the presence ofα,α-azoisobutyronitrile as a free radical initiator under refluxconditions for 16 hours can afford the monobromide as a major product(90 percent) and dibromide as a minor product (10 percent). Displacementof the bromide by formate followed by acid catalyzed hydrolysis to givehydroxymethyl derivative was carried out according to a modification ofa procedure described in Alpegiani, M. et al., Synth. Commun., 1992, 22,pp. 1277-1282.

The synthesis of carbamates from amines is well known. The inventionprovides for the synthesis of carbamates of aromatic amidines. Methylcarbamate 2 was synthesized by the known method of reacting bis-amidine1 with methylchloroformate in CH₂Cl₂ employing sodium hydroxide as abase, often obtaining yields less than 50 percent. An alternativeapproach to potentially improve the yield and purity of the carbamateinvolves reacting bis-amidine 1 with methyl-4-nitrophenyl carbonate inDMF to obtain methyl carbamate 2 at a yield of 80 percent. Carbamates3-10 set forth in Table 1 were synthesized from bis-amidine 1 and theappropriate carbonates in DMF or THF/CH₃CN (see Scheme 1). A base wasnot employed when using DMF. A base (e.g., diisopropylethylamine) wasused when employing THF/CH₃CN. Symmetrical carbonates were reacted withbis-amidine 1 to obtain the expected carbamates (7-9).

Table 1 contains the results of evaluation of the carbamate andcarbamate prodrugs 2-11 against P. carinii pneumonia in animmunosuppressed rat model (see S. K. Jones et al., Antimicrob. AgentsChemother. 1990, 34, pp. 1026-1030). Most of the prodrugs 2-11 appear tobe metabolized in vivo and were generally effective against PCP, orallyand intraveneously.

EXAMPLES

The invention will now be described in greater detail with reference tothe following examples. It should be noted that these examples are forillustrative purposes only, and are not meant to limit the invention.

Examples 1-25 Experimental Procedure

In the examples which follow, melting points were recorded using aThomas-Hoover (Uni-Melt) capillary melting point apparatus and wereuncorrected. TLC analysis was carried out on silica gel 60 F₂₅₄pre-coated aluminum sheets (0.20 mm layer thickness) (E. Merck ofWhitehouse Station, N.J.) and detected under UV light. IR spectra wererecorded using Perkin-Elmer Model 337 spectrometer sold by Perkin-Elmerof Norwalk, Conn. ¹H and ¹³C NMR spectra were recorded employing aVarian GX400 or a Varian Unityplus 300 spectrometer (both sold by Varianof Palo Alto, Calif.) and chemical shifts (δ) are in ppm relative to TMSas internal standard. Mass Spectra were recorded on a VG Analytical70-SE spectrometer (Georgia Institute of Technology, Atlanta, Ga.). IRspectra were recorded using a Perkin-Elmer 2000 instrument. Elementalanalyses were obtained from Atlantic Microlab Inc. of Norcross, Ga. andare believed to be within 0.4 percent of the theoretical values unlessotherwise mentioned. All chloroformates were purchased from AldrichChemical Co. of St. Louis, Mo. Other chemicals and solvents werepurchased from either Aldrich or Fischer Scientific of Houston, Tex.4,5-Dimethyl-1,3-dioxol-2-one was purchased from TCI America Inc.2,5-Bis(4-cyanophenyl)furan, 2,5-bis(4-amidinophenyl)furan, and2,5-bis[4-(N-hydroxy)amidinophenyl]furan were synthesized as previouslydescribed. See Baijic, M., et al., Heterocycl. Commun., 1996, 2,pp.135-140; Das, B. P., J. Med. Chem. 1977, 20, pp. 531-536; and Boykin,D. W., et al., Bioorg. Med. Chem Lett., 1996, 6, pp. 3017-3020.Anti-Pneumocystis carinii pneumonia activity screening was carried outaccording to published methods. See Jones, S. K. et al., Antimicrob.Agents Chemother., 1990, 34, pp.1026-1030; Hall, J. E., et al.,Antimicrob. Agents Chemother., 1998, 42, pp. 666-674; and Tidwell, R.R., et al., J. Med. Chem., 1990, 33, pp. 1252-1257. Compounds wereroutinely tested orally at 33 μmol/kg/day and intravenously at 22μmol/kg/day. Saline- and pentamidine treated groups of rats wereincluded as negative and positive controls, respectively.

Bold numbers that are listed in connection with each example correspondto the numbers listed in Table 1.

Example 1 Synthesis of Methyl 4-nitrophenyl Carbonate (12)

To an ice cold solution of 4-nitrophenol (7.36 g, 0.053 mol) andpyridine (4.3 g, 0.054 mol) in CH₂Cl₂ (80 mL) at 0-5° C. was added asolution of methylchloroformate(5 g, 0.053 mol) in CH₂Cl₂ (20 mL) andstirred for 15 min and then at room temperature overnight (16 h). Themixture was extracted with CH₂Cl₂ (50 mL), washed successively withwater (50 mL), aq. NaOH (0.5 N, 50 mL), sat. aq. NaCl solution (50 mL),water (3×50 mL) and dried (Na₂SO₄). The CH₂Cl₂ solution was passedthrough a silicagel column using chloroform (100%) as eluent to furnishpure carbonate 12 (10 g, 96%) as a white solid. Purification of thecarbonate by recrystallization gave 80% yield: TLC (R_(f)) 0.50 (100%CHCl₃); mp 111-112° C.; IR (KBr) 3121, 3086, 1766, 1618, 1602, 1522,1443, 1366, 938, 858, 667 cm⁻¹; ¹H NMR (CDCl₃) 8.24 (d, 2H, J=9.05 Hz,Ar—CH), 7.34 (d, 2H, J=9.05 Hz, Ar—CH), 3.91 (s, 3H, OCH₃); ¹³C NMR(CDCl₃), 155.71 (OCOO), 153.28 (Ar—OCO), 145.61 (C—NO₂), 125.51 (Ar—CH),121.96 (Ar—CH),56.07 (OCH₃); MS m/e (EI⁺, relative intensity, %) 197(25), 153 (33), 123 (100), 95 (21), 92 (45), 77 (46), 64 (32), 63 (33),59 (55).

Example 2 2,5-Bis[4-(N-methoxycarbonyl)amidinophenyl]furan (2)

To a stirring suspension of bis-amidine 1 (0.5 g, 0.00164 mol) in dryDMF (8 mL) at room temperature, was added a solution of methyl4-nitrophenyl carbonate (0.72 g, 0.0036 mol) in DMF (2 mL) and themixture was stirred overnight (16 h). Water (20 mL) was added to themixture, stirred for few min and filtered, washed with water (2×10 mL),and ether (10 mL) and dried. Crystallization from ethanol gave purecarbamate 2 (80%) as a white solid: TLC (R_(f)) 0.49 (CHCl₃, MeOH,NH₄OH, 4:1:0.2, v/v); mp>300° C. dec.; IR (KBr) 3500-3100, 3010, 2956,1672, 1609, 1566, 1518, 1476, 1442, 1266, 1198, 1147, 1124, 1086, 1030,940, 856, 806, 786, 764, 674, 604, 548 cm⁻¹; ¹H NMR (DMSO-d₆) δ 9.11 (s,4H, NH), 8.07 (d, 2H, J=8.18 Hz, Ar—CH), 7.95 (d, 2H, J=8.30 Hz, Ar—CH),7.30 (s, 2H, furan-CH), 3.63 (s, NCOOCH₃); ¹³C NMR (DMSO-d₆) δ 165.61,164.36, 152.61, 132.96, 132.81, 128.36, 123.34, 110.52, 51.85 MS m/z(FAB, thioglycerol) 421 (M+1), 389, 363, 346, 321, 305, 289, 271, 257,237, 230. Anal. (C₂₂H₂₀N₄O₅) C, H, N.

Example 3 Synthesis of 4-nitrophenyl-2,2,2-trichloroethyl Carbonate (14)

To an ice cold solution of 4-nitrophenol (2.0 g, 14.4 mmol) andtriethylamine (1.6 g, 15.8 mmol) (or pyridine) in CH₂Cl₂ (20 mL) at 0-5°C. was added a solution of 2,2,2-trichloroethylchloroformate (3.2 g, 15mmol) in CH₂Cl₂ (10 mL) and stirred for 15 min. and then at roomtemperature overnight (16 h). Aqueous work up as described above andpurification of the product by silicagel column chromatography usingchloroform (100%) as eluent furnished pure carbonate 14 (4.3 g 91%).Alternatively, the product is purified by recrystallization from hexanein 60% yield: TLC (R_(f)) 0.56 (100% CHCl₃); mp 59-60° C.; IR (KBr)3123, 3093, 3012, 2967, 2871, 2462, 2365, 2343, 1771, 1630, 1541, 1496,1432, 1362, 1288, 1251, 1020, 954, 843, 783 cm⁻¹; ¹H NMR (CDCl₃) δ 8.33(d, 2H, J=9.36 Hz, Ar—CH), 7.45 (d, 2H, J=9.36 Hz, Ar—CH), 4.91 (s, 2H,OCH₂CCl₃), ¹³C NMR (CDCl₃) δ 155.21 (OCOO), 151.68 (Ar—OCO), 145.91(C—NO₂), 125.64, 121.89, 93.95 (CCl₃), 77.56 (CH₂CCl₃); MS m/e (EI⁺,relative intensity, %) 314 (M⁺, 1), 313 (5), 280 (17), 278 (26), 196(14), 182 (25), 166 (74), 139 (100), 135 (20), 133 (58), 122 (24), 109(40), 95 (32), 63 (22). Anal. (C₉H₆NO₅Cl₃) C, H.

Example 4 Synthesis of2,5-bis[4-(N-2,2,2-trichloroethoxycarbonyl)amidinophenyl]furan (3)

To a suspension of amidine 1 (0.5 g, 0.00164 mol) anddiisopropylethylamine (0.43 g, 0.0033 mol) in THF/CH₃CN mixture (20 mL,1:1 v/v) at room temperature, was added a solution of4-nitrophenyl-2,2,2-trichloroethyl carbonate (1.1 g, 0.0035 mol) in THF(10 mL) and stirred for 24 h. The solvents were removed in a rotavap andthe residue was cooled in ice, triturated with anhydrous diethyl ether(20 mL), filtered, washed with ether (2×20 mL), dried and crystallizedfrom CHCl₃-ether mixture to obtain 2,2,2-trichloroethylcarbamate (3)(0.65 g, 60% yield) as a yellow solid: TLC (R_(f)) 0.6; (CHCl₃, MeOH,NH₄OH, 4:1:0.2, v/v); mp 134-136° C.; IR (KBr) 3509-3029, 3010, 2997,2952, 1682, 1615, 1600, 1517, 1492, 1485, 1412, 1377, 1251, 1147, 1130,1120, 1058, 1028, 939, 849, 798, 730, 716, 634, 560 cm⁻¹; ¹H NMR(DMSO-d₆) δ 9.80-9.60 (br s, 4H, NH), 8.07 (d, J=8.73 Hz, 4H, Ar—CH),8.02 (d, J=8.73 Hz, 4H, Ar—CH), 7.39 (s, 2H, CH-furan), 4.98 (s, 4H,OCH₂CCl₃); ¹³C NMR (DMSO-d₆) δ 166.35, 152.58, 133.57, 129.23, 123.39,111.23, 95.57 (CCl₃), 74.49 (CH₂CCl₃); MS m/z (FAB, thioglycerol): 656(M+1, 9 isotopic peaks), 507 (7 peaks), 481 (8 peaks), 481 (8 peaks),464.0 (6 peaks), 357 (3 peaks), 314 (3 peaks), 304, 288, 271, 262, 245,232. Anal. (C₂₄H₁₈N₄O₅Cl₆) C, H, N.

Example 5 Synthesis of 4-nitrophenyl thioethyl Carbonate (18)

Thiocarbonate 18 was synthesized from 4-nitrophenol andthioethylchloroformate in pyridine/CH₂Cl₂ as described previously, in92% yield and subsequently purified by silica column chromatography toafford pure colorless crystals: TLC (R_(f)) 0.6 (100% CHCl₃); mp 65-66°C.; IR (KBr) 3120, 3089, 2944, 2588, 2000, 1942, 1740, 1528, 1454, 1352,1198, 1102, 894, 748, 660, 526 cm⁻¹; ¹H NMR (CDCl₃) d 8.25 (dd, 2H,J=4.92, 2.07 Hz, Ar—CH), 7.33 (dd, 2H, J=4.92, 2.22 Hz, Ar—CH), 2.96 (q,2H, J=14.84 Hz, SCH₂), 1.38 (t, 3H, J=7.46 Hz, CH₃); ¹³C NMR (CDCl₃) d169.70, 155.79, 145.52, 125.44, 122.21, 26.04 (SCH₂), 14.85 (CH₃); MSm/e (EI⁺, relative intensity, %) 227 (M⁺, 4), 139 (12), 109 (19), 89(100), 63 (12). Anal. (C₉H₉NO₄S) C, H.

Example 6 Synthesis of2,5-bis[4-(N-thioethylcarbonyl)amidinophenyl]furan(4)

To a suspension of bis-amidine 1 (0.6 g, 0.002 mol) in DMF (10 mL) atroom temperature, was added a solution of 4-nitrophenylethylthiocarbonate (0.9 g, 0.004 mol). The resulting solution wasstirred for 24 h. Ice water (40 mL) was added to the mixture andfiltered, washed with water (3×30 mL), ether (30 mL) and dried. Thecrude solid was purified by crystallization from ethanol-ether tofurnish carbamate 4 (0.6 g, 62%) as a yellow solid: TLC (R_(f)) 0.58(CHCl₃, MeOH, NH₄OH, 4:1:0.2, v/v); mp>300°; IR (KBr) 3442-3200, 3040,2970, 2928, 2866, 1668, 1610, 1592, 1562, 1469, 1413, 1382, 1320, 1305,1283, 1208, 1130, 1110, 1090, 1016, 939, 885, 842, 768 cm⁻¹; ¹H NMR(DMSO-d₆) δ 9.26 (d, 4H, D₂O exchangeable, J=45 Hz, NH), 8.07 (d, 4H,J=8.42 Hz, Ar—CH), 7.96 (d, 4H, J=8.30 Hz, Ar—CH), 7.31 (s, 2H,CH-furan), 2.78 (q, 4H, J=14.3 Hz, SCH₂), 1.24 (t, 6H, J=7.32 Hz,SCH₂CH₃); ¹³C NMR (DMSO-d₆)_(+ δ) 181.37, 162.06, 152.64, 132.95,132.35, 128.70, 123.43, 110.76, 24.06 (SCH₂), 15.34 (SCH₂CH₃); MS m/z(FAB, m-nitrobenzoic acid) 481 (M+1), 429, 413, 397, 321, 298, 272, 257,231. Anal. (C₂₄H₂₄N₄O₃S₂.0.25H₂O) C, H, N.

Example 7 Benzyl-4-nitrophenylcarbonate (19)

Carbonate 19 was synthesized from 4-nitrophenol and benzylchloroformateas described above, in 80% yield after silica column purification, as awhite solid: TLC (R_(f)) 0.55 (100% CHCl₃); mp 81° C.; IR (KBr) 3098,3034, 2855, 1758, 1617, 1528, 1387, 1355, 1285, 1227, 1112, 1049, 965,870, 780, 729, 583, 511 cm⁻¹; ¹H NMR (CDCl₃) d 8.26 (d, 2H, J=9.4 Hz),7.43 (m, 5H, Ar—CH of benzyl), 7.37 (d, 2H, J=9.4 Hz, OCH₂—Ar); ¹³C NMR(CDCl₃): δ 155.53, 152.45, 145.40, 134.20, 129.07, 128.81, 128.66,125.30, 121.77, 71.01; MS m/e (EI⁺, relative intensity, %) 274, 244,230, 199, 140, 131, 108.

Example 8 Synthesis of2,5-bis[4-(N-benzyloxycarbonyl)amidinophenyl]furan (5) and DimaleateSalt

To a suspension of bis-amidine 1 (0.5 g, 0.0016 mol) in DMF (10 mL) atroom temperature, was added benzyl-4-nitrophenylcarbonate (1.6 g, 0.006mol). The resulting solution was stirred for 24 h and ice water (40 mL)was then added and extracted with CHCl₃ (2×50 mL). The CHCl₃ extract waswashed with aq. NaOH (1 N, 40 mL), sat. NaCl (40 mL), water (50 mL) anddried (Na₂SO₄). The solution was filtered, concentrated in a rotavap,cooled in ice bath, triturated with ether (30 mL), filtered, washed withether (3×20 mL) and dried under vacuum for 16 h to afford carbamate 5 asa shiny pale yellow solid (0.77 g, 52%): TLC (R_(f)) 0.76 (CHCl₃, MeOH,NH₄OH, 4:1:0.2, v/v); mp 225° C. dec.; IR (KBr) 3480-3140, 3111, 3087,3063, 3032, 2960, 2866, 1667, 1612, 1570, 1507, 1497, 1375, 1296, 1266,1145, 926, 859, 787, 744, 702 cm⁻¹; ¹H NMR (DMSO-d₆) δ 9.16 (s, 4H, D₂Oexchangeable, NH), 8.09 (d, 4H, J=8.57 Hz, Ar—CH), 7.95 (d, 4H, J=8.57Hz, Ar—CH), 7.42-7.31 (2×d+3×t, 10H, Ar—CH), 7.30 (s, 2H, CH-furan),5.13 (s, OCH₂Ph, 4H ¹³C NMR (DMSO-d₆) δ 165.97, 163.69, 152.62, 137.12,132.89, 132.86, 128.44, 128.35, 127.97, 127.77, 123.35, 110.60, 66.04(OCH₂Ph); MS m/e (FAB, m-nitrobenzoic acid) 573 (M+1), 460, 439, 421,378. Anal. (C₃₄H₂₈N₄O₅.2H₂O) C, H, N.

A mixture of the carbamate free base 5 (0.4 g, 0.0007 mol), maleic acid(0.18 g, 0.0016 mol) and dry ethanol (20 mL) was stirred at roomtemperature for 4 h. The mixture was cooled in ice bath, triturated withdry Et₂O (25 mL), filtered, washed with Et₂O (3×10 mL) and dried in avacuum oven at 50° C. overnight to afford 5-dimaleate salt as a yellowsolid (0.53 g, 94%): mp 155-157° C. dec.; IR (KBr); 3480-3140, 3111,3087, 3063, 3032, 2960, 1667, 1612, 1570, 1507, 1497, 1375, 1266, 1145,926, 859, 787, 744, 702 cm⁻¹; ¹H NMR (DMSO-d₆) δ 9.40-9.80 (br s, 2H,D₂O exchangeable, NH), 8.03 (d, 4H, J=8.56 Hz, Ar—CH), 7.99 (d, 4H,J=8.56 Hz, Ar—CH), 7.42-7.31 (2×d+3×t, 10H, Ar—CH), 7.35 (s, 2H,CH-furan), 6.18 (s, 2H, O₂CCH=CHCO₂), 4.19 (s, 4H, OCH₂Ph); ¹³C NMR(DMSO-d₆) δ 166.77, 165.49, 152.57, 136.39, 133.34, 132.07, 128.94,128.39, 128.11, 128.01, 123.36, 111.03, 66.72 (OCH₂Ph); MS m/e (FAB,m-nitrobenzoic acid) (free base) 573.2, 460.1, 421.2, 378.2; Anal.(C₄₂H₃₆N₄O₁₃) C, H, N.

Example 9 Synthesis of 4-bromomethyl-5-methyl-1,3-dioxol-2-one

A mixture of 4,5-dimethyl-1,3-dioxol-2-one (15.0 g, 0.132 mol),a,a-azobisisobutyronitrile (AIBN) (1.08 g, 0.0066 mol) andN-bromosuccinimide (23.4 g, 0.132 mol) in freshly distilled carbontetrachloride (350 mL) was refluxed under nitrogen for 16 h. The mixturewas concentrated to one-half the initial volume, cooled in an ice bathand the white solid was filtered off. Concentration of the filtrate(CCl₄ solution) in a rotavap under reduced pressure gave4-bromomethyl-5-methyl-1,3-dioxol-2-one as a pale brown liquid in 90%yield (25 g). Due to the instability of the product at room temperature,it was used without purification for the next step. However, a smallamount of the crude produce (100 mg) was purified for NMR analysis,through a short pad of silicagel using CHCl₃ (100%) as eluent. ¹H NMR(CDCl₃) δ 4.21 (s, 2H, CH₂Br), 2.14 (s, 3H, CH₃); ¹³C NMR (CDCl₃) δ151.72 (OCOO, 138.08 (═CCH₂Br), 134.62 (═C—CH₃), 18.01 (CH₂Br), 9.51(CH₃).

Example 10 Synthesis of (5-methyl-1,3-dioxol-2-one-4-yl)methyl formate

To an ice cold solution of 4-bromomethyl-5-methyl-1,3-dioxol-2-one (24g, 0.124 mol) and formic acid (19.5 g, 0.43 mol) in acetonitrile (250mL) at 0° C. was added triethylamine (44 g, 0.44 mol) dropwise over aperiod of 15 min. Ice bath was then removed and the mixture was stirredat room temperature for 2 h. The mixture was concentrated to one-halfthe initial volume on a rotavap and extracted with ethyl acetate (2×150mL). The organic extract was washed successively with satd. NaHCO₃ (200mL), satd. NaCl (200 mL), water (200 mL) and dried (Na₂SO₄).Concentration of the filtrate gave crude formate ester as a colorlessliquid (20 g): ¹H NMR (CDCl₃) δ 8.06 (s, 1H, CH₂OOCH), 4.91 (s, 2H,CH₂), 2.16 (s, CH₃); ¹³C NMR (CDCl₃) δ 160.26 (CH₂OOCH), 152.06 (OCOO),140.59 (═CCH₂), 133.14 (═C—CH₃), 53.15 (CH₂OOCH), 9.42 (CH₃).

Example 11 Synthesis of 4-Hydroxymethyl-5-methyl-1,3-dioxol-2-one

To a solution of the crude formate (19.9 g) and 80% methanol (250 mL) atroom temperature was added conc. HCl (1 mL) and stirred for 6 h.Methanol was evaporated off in rotavap at 30° C. under reduced pressureand the residue was extracted with ethyl acetate. Passage through ashort pad of silica gel and concentration gave4-hydroxymethyl-5-methyl-1,3-dioxol-2-one as a colorless oil (7.0 g,43%): ¹H NMR (CDCl₃) δ 4.37 (s, 2H, CH₂OH), 2.80 (s, 1H, OH, D₂Oexchangeable), 2.11 (s, 3H, CH₃); ¹³C NMR (CDCl₃) δ 152.94 (OCOO),137.63 (═CCH2OH), 135.01 (═CCH₃), 53.32 (CH₂OH), 9.33 (CH₃).

Example 12 Synthesis of4-methyl-2-oxo-1,3-dioxiol-4-en-5-yl)methyl-4-nitrophenyl carbonate (23)

Carbonate 23 was synthesized from4-methyl-5-hydroxymethyl-1,3-dioxol-4-ene-2-one and4-nitrophenylchloroformate in CH₂Cl₂/pyridine and purified bycrystallization from chloroform colorless crystals in 76% yield: TLC(R_(f)) 0.23 (100% CHCl₃); mp 120-121° C. (lit.³¹ 116-117° C.); IR (KBr)3115, 3096, 2928, 2854, 1811, 1780, 1742, 1619, 1593, 1525, 1494, 1352,1308, 1246, 1221, 1054, 860, 768 cm⁻¹; ¹H NMR (CDCl₃) δ 8.29 (d, 2H,J=9.04 Hz), 7.39 (d, 2H, J=9.20 Hz), 5.03 (s, 3H, C═CCH₃), 2.22 (s, 2H,OCOOCH₂); ¹³C NMR (CDCl₃) δ 155.36 (OC═OO), 152.46, 151.85 (vinyleneC═O), 145.93 (Ar—NO₂), 141.62, 132.45, 125.59, 121.90, 58.37 (OCH₂),9.65 (C═CCH₃); MS m/e (EI⁺, relative intensity, %) 295 (M⁺, 1), 139 (9),113 (100), 69 (23), 43 (74). Anal. (C₁₂H₉NO₈) C, H, N.

Example 13 Synthesis of2,5-bis{4-[N-(5-methyl-2-oxo-1,3-dioxol-4-ene-1-yl)methoxycarbonyl]amidinophenyl}furan (6)

To a suspension of bis-amidine 1 (0.7 g, 0.0023 mol) in DMF (15 mL) atroom temperature, was added a solution of(4-methyl-2-oxo-1,3-dioxol-4-ene-1-yl) methyl 4-nitrophenyl carbonate(1.5 g, 0.0052 mol) in DMF (5 mL) and stirred for 24 h. Ice water (50mL) was added, filtered, washed with water (3×20 mL), ether (30 mL) anddried under vacuum. The crude product was crystallized from CHCl₃-ethermixture to yield pure 6 (1.26 g. 89%) as a yellow solid: TLC (R_(f))0.33 (CHCl₃, MeOH, NH₄OH, 4:1:0.2, v/v); mp 153-155° C.; IR (KBr)3500-3120, 3105, 3075, 2866, 3037, 1825, 1667, 1660, 1618, 1610, 1521,1497, 1417, 1394, 1266, 1230, 1145, 1094, 989, 927, 787, 687 cm⁻¹; ¹HNMR (DMSO-d₆) δ 9.19 (s, 4H, D₂O exchangeable, NH), 8.09 (d, J=8.43 Hz,Ar—CH), 7.96 (d, 4H, J=8.24 Hz, Ar—CH), 7.31 (s, 2H, CH-furan), 4.95 (s,4H, OCH₂), 2.18 (s, 6H, C═CCH₃); ¹³C NMR (DMSO-d₆) δ 166.23, 162.91,152.60 (C═O, dioxolone), 151.96, 139.53, 134.09, 132.96, 132.65, 128.47,123.35, 110.66, 54.17 (OCH₂), 8.82 (C═CCH₃); MS m/z (FAB, m-nitrobenzoicacid) 617 (M+1), 505, 487, 460, 443, 424, 375, 357. Anal. (C₃₆H₂₄N₄O₁₁)C, H, N.

Example 14 Synthesis of Diphenyl Carbonate (20)

Carbonate 20 was prepared by reaction of phenol withphenylchloro-formate in pyridine/CH₂Cl₂ followed by aqueous workup asdescribed above in 90% yield as a white solid: TLC (R_(f)) 0.7 (100%CHCl₃); mp 79-80° C.; IR (KBr) 3066, 1776, 1604, 1495, 1457, 1400, 1285,1189, 997, 755, 691 cm⁻¹; ¹H NMR (CDCl₃) δ 7.40 (t, 4H, J=8.02 Hz), 7.26(d, 6H, J=8.73 Hz); ¹³C NMR (CDCl₃) δ 152.29, 151.14, 129.77, 126.50,121.10; MS m/e (EI⁺, relative intensity, %) 214 (M⁺, 100), 170 (46), 169(37), 142 (43), 141 (64), 94 (13), 77 (87), 65 (26), 51 (25), 39 (23).

Example 15 Synthesis of 2,5-Bis[4-(N-phenoxycarbonyl)amidinophenyl]furan(7)

To a suspension of bis-amidine 1 (0.5 g, 0.0016 mol) in DMF (10 mL) atroom temperature, was added diphenylcarbonate (0.77 g, 0.0036 mol). Theresulting solution was stirred for 24 h and ice water (40 mL) was thenadded and the resulting solid was filtered, washed with plenty of water(3×30 mL), ether (2×30 mL) and dried under vacuum in a dessicator for 16h to furnish carbamate 7 (0.53 g, 63%) as a yellow solid: TLC (R_(f))0.68 (CHCl₃, MeOH, NH₄OH, 4:1:0.2, v/v); mp>300° C.; IR (KBr) 3680-3000,1674, 1615, 1462, 1515, 1488, 1412, 1382, 1266, 1199, 1170, 1145, 1030,1016, 939, 969, 864, 798, 738, 693, 589 cm⁻¹; ¹H NMR (DMSO-d₆) δ 9.30(s, 4H, D₂O exchangeable, NH), 8.12 (d, 4H, J=7.61 Hz, Ar—CH), 7.98 (d,4H, J=7.30 Hz, Ar—CH), 7.44 (t, 4H, J=7.46 Hz, Ar-m-H), 7.33 (s, 2H,CH-furan), 7.22 (t, 2H, J=6.19 Hz, Ar-p-CH), 7.20 (d, 4H, J=8.57 Hz,Ar-o-CH); ¹³C NMR (DMSO-d₆) δ 166.78, 162.12, 152.65, 151.63, 151.63,133.08, 132.58, δ129.17, 128.59, 124.92, 123.43, 121.95, 115.19, 110.79;MS m/z (FAB, m-nitrobenzoic acid) 545 (M+1), 460, 451, 425, 408, 391,357, 329. Anal. (C₃₂H₂₄N₄O₅) C, H, N.

Example 16 Synthesis of bis(4-flourophenyl)carbonate (21)

Reaction of 4-fluorophenol with 4-fluorophenylchloroformate inpyridine/CH₂Cl₂ as described earlier afforded carbonate 21 after silicacolumn chromatography in 85% yield as a white solid: TLC (100% CHCl₃)0.7; mp 122-123° C.; IR (KBr) 3130, 3091, 1885, 1764, 1649, 1610, 1508,1304, 1234, 1176, 1094, 1010, 902, 838, 729, 576, 510 cm⁻¹; ¹H NMR(CDCl₃) δ 7.24 (dd, 4H, J=9.05, 4.44 Hz), 7.08 (dd, 4H, J=8.89, 8.09Hz); ¹³C NMR (CDCl₃) δ 161.93, 159.50, 152.34, 147.06, 147.03, 122.61,122.53, 116.60, 116.36; MS m/e (EI⁺, relative intensity, %) 250 (M⁺, 82)206 (27), 178 (12), 177 (43), 139 (11), 112 (25), 111 (20), 95 (100), 83(32), 75 (19), 57 (17).

Example 17 Synthesis of2,5-bis[4-(N-(4-fluoro)phenoxy-carbonyl)amidinophenyl]furan (8)

To a suspension of bis-amidine 1 (0.5 g, 0.0026 mol) in DMF (10 mL) atroom temperature, was added a solution of carbonate 21 (0.87 g, 0.0035mol). The resulting solution was stirred for 16 h. ice water (40 mL) wasadded to the mixture and filtered, washed with water (3×30 mL), ether(30 mL) and dried in vacuum for 24 h to furnish 4-fluorophenylcarbanate8 (0.92 g, 61%) as a yellow solid: TLC (R_(f)) 0.45 (CHCl₃, MeOH, NH₄OH,4:1:0.2, v/v); mp>300° C.; IR (KBr) 3465-3000, 1667, 1621, 1491, 1260,1187, 1139, 1078, 969, 859, 793, 665 cm⁻¹; ¹H NMR (DMSO-d₆) δ 9.31 (s,4H, D₂O exchangeable, NH), 8.12 (d, J=8.73 Hz, Ar—CH), 7.98 (d, 4H,J=8.57 Hz, Ar—CH), 7.33 (s, 2H, CH-furan), 7.22 (d, 8H, J=6.5 Hz,F—Ar—CH) ¹³C NMR (DMSO-d₆) δ 162.12, 161.86, 157.77, 156.53, 154.21,153.45, 152.57, 147.76, 133.00, 132.45, 132.65, 128.79, 128.44, 127.83,123.46, 123.38, 123.31, 115.97, 115.89, 115.60, 115.47, 115.37, 115.25;MS m/z (FAB, m-nitrobenzoic acid) 581 (M+1), 469, 443, 426, 357, 331.Anal. (C₃₂H₂₂N₄O₅F₂.0.5H₂O) C, H, N.

Example 18 Synthesis of bis(4-methoxyphenyl)carbonate (22)

Reaction of 4-methoxyphenol with 4-methoxyphenylchloroformate inpyridine/CH₂Cl₂ followed by aqueous workup as described above gavecarbonate 22, after silica column chromatography, in 93% yield as awhite solid: TLC (R_(f)) 0.53 (100% CHCl₃); mp 95° C.; IR (KBr) 3076,2958, 2848, 1772, 1610, 1514, 1470, 1286, 1242, 1182, 1028, 894, 836,776, 726, 534 cm⁻¹; ¹H NMR (CDCl₃) δ 7.16 (d, 4H, J=9.05 Hz), 6.88(d,4H, J=9.04 Hz), ¹³C NMR (CDCl₃) δ 157.71, 153.04, 144.83, 121.96,114.68, 55.80 (OCH₃); MS m/e (EI⁺, relative intensity, %) 274 (M⁺, 100),230 (33), 215 (29), 187 (12), 124 (16), 123 (46), 107 (10), 95 (12), 77(13), 64 (7), 52 (5), 41 (6).

Example 19 Synthesis of 2,5-bis[4-(N-(4-methoxy)phenoxycarbonyl)amidinophenyl]furan (9)

To a suspension of bis-amidine 1 (0.7 g, 0.0016 mol) in DMF (10 mL) atroom temperature, was added bis(4-methoxy)phenylcarbonate (1.39 g,0.0051 mol) and stirred for 24 h. Anhydrous ether (25 mL) was then addedto the precipitated product, stirred for few min and filtered, washedwith ether (3×15 mL) and dried under vacuum in a dessicator for 48 h tofurnish 4-methoxyphenyl carbamate 9 (0.9 g, 65%) as a yellow solid: TLC(R_(f)) 0.68 (CHCl₃,MeOH, NH₄OH, 4:1:0.2, v/v); mp>300° C.; IR (KBr)3450-3100, 3010, 2934, 2836, 1683, 1484, 1256, 1184, 1142, 1078, 1033,1010, 967, 928, 850, 801, 774, 753, 696, 659, 607, 583, 559, 531 cm⁻¹;¹H NMR (DMSO-d₆) δ 9.26 (s, 4H, NH), 8.11 (d, 4H, J=8.54 Hz, Ar—CH),7.98 (d, 4H, J=8.53 Hz, Ar—CH), 7.34 (s, 2H, furan-CH), 7.09 (d, 4H,J=9.04 Hz, Ar—CH of Ar—OCH₃), 6.93 (d, 4H, J=9.03 Hz, Ar—CH of Ar—OCH₃),3.75 (s, 6H, OCH₃); ¹³C NMR (DMSO-d₆) δ 166.62, 156.26, 152.64, 145.07,133.03, 132.64, 128.53, 123.41, 122.66, 114.12, 110.72, 55.36. MS m/z(FAB, thioglycerol) 605 (M+1), 481, 429, 323, 303, 289, 273, 257, 247,229. Anal. (C₃₄H₂₈N₄O₇.1.DMF) C, H, N.

Example 20 Synthesis of 1-chloroethyl-4-nitrophenylcarbonate (16)

To an ice cold solution of 4-nitrophenol (2.0 g, 0.015 mol) andtriethylamine (1.6 g, 0.016 mol) (or pyridine) in CH₂Cl₂ (20 mL) at 0-5°C. was added a solution of 1-chloroethylchloroformate (2.1 g, mmol) inCH₂Cl₂ (10 mL) and stirred for 15 min and then at room temperatureovernight (16 h). The mixture was extracted with CH₂Cl₂ (50 mL), aq.NaOH (0.5 N. 50 mL), sat. NaCl solution (50 mL), water (3×50 mL) anddried (Na₂SO₄). The CH₂Cl₂ solution was filtered, evaporated in arotavap and the residue was purified by silicagel column chromatographyusing chloroform (100%) as eluent to furnish pure 16 as a white solid:TLC (R_(f)) 0.75 (CHCl₃); mp 70-71° C. (lit.²² 69-70° C.); IR (KBr)3116, 3084, 2999, 2932, 2864, 2364, 2330, 1779, 1626, 1525, 1355, 1245,1101, 914, 863, 779, 677 cm⁻¹; ¹H NMR (CDCl₃) δ 8.31 (dd, 2H, J=5.08,2.07 Hz, Ar—CH), 7.43 (dd, 2H, J=4.76, 2.22 Hz, Ar—CH), 6.50 (q, 1H,J=11.67 Hz, CHClCH₃), 1.93 (d, 3H, J=5.87 Hz); ¹³C NMR (CDCl₃) δ 155.16,150.65, 145.94, 125.63, 121.89, 85.44, 25.34; MS m/e (EI⁺, relativeintensity) 210 (M⁺—HCl, 4), 139 (26), 122 (13), 109 (8), 76 (13), 75(11), 65 (27), 64 (17), 63 (100), 50 (10), 43 (13).

Example 21 Synthesis of 1-acetoxyethyl-4-nitrophenylcarbonate (17)

To a solution of 1-chloroethyl-4-nitrophenyl carbonate (2.0 g, 0.0082mol) in glacial acetic acid (50 mL) at room temperature, was addedmercuric acetate (3.8 g, 0.012 m) and the mixture was stirred for 40 h.Water (100 mL) was then added to the mixture and extracted with ether(2×75 mL). The ethereal phase was washed with aq. NaOH (0.5 N, 30 mL),sat. NaCl (30 mL), water (2×50 mL) and dried (anhy. Na₂SO₄). Thesolution was filtered, concentrated in a rotavap and purified by silicagel column chromatography to afford pure 1-acetoxyethyl-4-nitrophenylcarbonate (17) (1.9 g, 89%) as a colorless liquid: TLC (R_(f)) 0.65(CHCl₃); IR (film) 1779 (OCOO), 1749 (CH₃COO), 1615, 1592, 1528, 1491,1266, 1110, 1070, 857 cm⁻¹; ¹H NMR (CDCl₃) δ 8.29 (d, 2H, J=9.05 Hz,Ar—CH), 7.41 (d, 2H, J=9.04 Hz, Ar—CH), 6.84 (q, 1H, J=10.95 Hz,CH(Oac)CH₃), 2.14 (s, 3H, COCH₃), 1.62 (d, 3H, 5.4 Hz, CHCH₃); ¹³C NMR(CDCl₃) δ 169.09, 155.33, 150.70, 145.76, 125.52, 121.94, 92.47 (CHOAc),20.96 (COCH₃), 19.61 (CHCH₃); MS m/e (EI⁺, relative intensity) 210(M⁺—AcOH, 3), 166 (4), 122 (5), 87 (33), 63 (6), 50 (3), 43 (100).

Example 22 Synthesis of2,5-bis[4(1-acetoxyethoxycarbonyl)amidinophenyl]furan (10)

A mixture of bis-amidine 1 (0.4 g, 0.0013 mol), diisopropylethylamine(0.35 g, 0.0026 mol) and THF/CH₃CN (1:1 mixture, 15 mL) was stirred atroom temperature. A solution of 1-aceoxyethyl-4-nitrophenyl carbonate(0.71 g, 0.00264 mol) in THF (5 mL) was then added and continuedstirring for 24 h. Solvents were removed in a rotavap under reducedpressure at 40° C., triturated with anhy. ether (20 mL), filtered,washed with ether (2×25 mL), dried in air and crystallized fromCHCl₃ether to yield 1-acetoxyethyl carbamate (10) as a yellow solid in71% yield (0.52 g): TLC (R_(f)) 0.5 (CHCl₃,MeOH, NH₄OH, 4:1:0.2, v/v);mp 165-167° C. dec; IR (KBr) 3690-2900 (br), 3458 (s), 3324 (s), 3131(s), 2945 (s), 1734, 1667, 1640, 1607, 1562, 1488, 1412, 1362, 1279,1243, 1147, 1117, 1089, 1057, 1022, 992, 932, 885, 842, 797, 597, 566cm⁻¹; ¹H NMR (DMSO-d₆) δ 9.33 (s, 4H, NH), 8.09 (d, 4H, J=8.54 Hz,Ar—CH), 7.96 (d, 4H, J=8.54 Hz, Ar—CH), 7.34 (s, 2H, CH-furan), 6.79 (q,2H, J=10.87 Hz, CHOAc), 2.03 (s, 6H, CH₃), 1.55 (d, 6H, J=5.39 Hz,CHCH₃); ¹³C NMR (DMSO-d₆) δ 168.93, 166.84, 161.49, 152.65, 133.06,132.58, 128.59, 123.42, 110.82, 89.18, 20.82, 19.63 (CH₃); MS m/z (FAB,thioglycerol) 565 (M+1), 479, 461, 435, 375, 357, 331, 314, 288, 271.Anal. (C₂₈H₂₈N₄O₉) C, H, N.

Example 23 Synthesis of2,5-bis[4(N-ethoxycarbonyloxy)amidinophenyl]furan (11): NaOH method

To a suspension of the bis-amidoxime (2,5-bis[4-(N-hydroxy)amidinophenyl]furan) (0.86 g, 0.0028 mol) and CH₂Cl₂ (15 mL), a solution ofethylchloroformate (1.22 g, 0.011 mol) in CH₂Cl₂ (15 mL) was added andstirred for 10 min. Aq. NaOH (1 N, 12 mL) was then added dropwise andstirred at room temperature for 6 h. Ice water (10 mL) was added,filtered, washed with plenty of water (3×30 mL), dried in air andcrystallized from ethanol to give pure ethyl carbonate (11) (0.67 g, 50%yield) as a white solid.

Example 24 Synthesis of ethyl 4-nitrophenylcarbonate (13):CarbonateMethod

Reaction of 4-nitrophenol with ethylchloroformate in pyridine/CH₂Cl₂ asdescribed earlier, gave carbonate 2 as colorless crystals in 92% yieldby chromatographic purification and 82% by crystallization methods. TLC(R_(f)) 0.48 (100% CHCl₃); mp 70-71° C.; IR (KBr) 3124, 3092, 3010,2920, 2866, 1772, 1622, 1600, 1536, 1278, 1112, 1060, 1006, 908, 860,774, 732, 662, 527, 502 cm⁻¹; ¹H NMR (CDCl₃) δ 8.29 (d, 2H, J=9.05 Hz,Ar—CH), 7.38 (d, 2H, J=9.05 Hz, Ar—CH), 4.36 (q, 2H, OCH₂, J=14.28 Hz),1.38 (t, 3H, J=7.07 Hz, CH₃): ¹³C NMR (CDCl₃) δ 155.84, 152.65, 145.64,125.48, 121.98, 65.74, 14.35; MS m/e (EI⁺, relative intensity, %) 212(M⁺, 1.4), 211 (1), 139 (100), 109 (60), 89 (100), 93 (13), 81 (11), 65(21), 63 (13).

Reaction of bis-amidoxime with ethyl 4-nitrophenyl carbonate in DMF atroom temperature gave bis-ethoxycarbonyloxy derivative in 85% yield as awhite solid. The physical data for compound 11 obtained by both methodswere virtually identical. TLC (R_(f)) 0.5 (CHCl₃, MeOH, NH₄OH, 4:1:0.2,v/v); mp>300° C. dec.; IR (KBr) 3700-3100, 3056, 2989, 2937, 2915, 2890,1770, 1668, 1635, 1481, 1414, 1370, 1266, 1208, 1124, 1035, 1013, 939,857, 834, 775, 686 cm⁻¹; ¹H NMR (DMSO-d₆) δ 7.91 (d, 4H, J=7.21 Hz,Ar—CH), 7.78 (d, 4H, J=7.20 Hz, Ar—CH), 7.24 (s, 2H, CH-furan), 6.89 (s,4H, NH), 4.20 (q, 4H, J=14 Hz, NOCOOCH₂), 1.26 (t, 6H, J=7.1 Hz, CH₃);¹³C NMR (DMSO-d₆) δ 156.0 (OCOO), 153.48, 152.43, 131.63, 130.17,127.29, 123.33, 109.70, 63.56, (OCH₂), 14.20 (OCH₂CH₃); MS m/z (FAB,thioglycerol) 481 (M+1), 429, 393, 377, 347, 323, 305, 288, 271, 237.Anal. (C₂₄H₂₄N₄O₇) C, H, N.

In the specification, and examples there have been disclosed typicalpreferred embodiments of the invention and, although specific terms areemployed, they are used in a generic and descriptive sense only and notfor the purposes of limitation, the scope of the invention being setforth in the following claims.

TABLE 1 In vivo Activity of Carbamate and Carbonate Prodrugs of2,5-Bis(4-amidinophenyl)furan vs. Pneumocystis carinii

Dosage^(a) Cysts/g of lung^(a) Compound R (μmol/kg/day) (% of control)Toxicity^(a) Saline — 100.0^(b) ± 13.24  0 Pentamidine iv @ 22.0 3.06 ±0.90 ++  1 H iv @ 13.3 0.83 ± 0.36 0 Oral @ 39.8 44.52 ± 13.30 0  2^(c)

iv @ 22.0 Oral @ 33.0 6.91 ± 6.01 49.68 ± 20.50 0 0  3^(d)

iv @ 22.0 Oral @ 33.0 1.85 ± 1.79 8.59 ± 9.14 0 0  4

iv @ 22.0 Oral @ 33.0 83.01 ± 43.65 19.52 ± 14.22 0 0  5^(c)

iv @ 11.0 Oral @ 33.0 0.03 ± 0.02 18.09 ± 9.16  0 0  6^(d)

iv @ 22.0 Oral @ 33.0 0.02 ± 0.01 18.73 ± 11.87 0 0  7

iv @ 22.0 Oral @ 33.0 3.61 ± 1.80 5.70 ± 5.15 0 0  8^(d)

iv @ 22.0 Oral @ 33.0 0.02 ± 0.01 2.21 ± 0.33 0 0  9^(d)

iv @ 22.0 Oral @ 33.0 0.02 ± 0.01 2.10 ± 2.08 0 0 10^(c)

iv @ 11.0 Oral @ 33.0 1.21 ± 1.02 57.16 ± 10.19 ++ 0 11^(c)

iv @ 34.7 Oral @ 33.0 1.66 ± 0.58 96.90 ± 48.48 0 0

Elemental Anaysis Data Calcd Calcd Calcd Found Found Found Compound forfor for for for for # Formula C H N C H N 2 C₂₀H₂₀N₄O₅ 62.84 4.80 13.3363.01 4.74 13.20 3 C₂₄H₁₈N₄O₅Cl₁₆ 41.70 3.20 8.11 41.54 3.01 8.28 4C₂₄H₂₄N₄O₃S₂.0.25H₂O 59.42 5.09 11.55 59.26 4.98 11.29 5 C₃₄H₂₈N₄O₅.2H₂O67.09 5.30 9.21 66.67 4.99 9.24   5M C₃₄H₂₈N₄O₅.2C₄H₄O₄ 62.68 4.51 6.9662.71 4.47 7.04 6 C₃₀H₂₄N₄O₁₁ 58.44 3.92 9.09 58.40 4.00 9.09 7C₃₂H₂₄N₄O₅.1.3H₂O 67.67 4.72 9.87 67.29 4.53 10.35 8 C₃₂H₂₂N₄O₅F₂.0.5H₂O65.12 4.10 9.51 64.82 4.20 10.03 9 C₃₄H₂₈N₄O₇.DMF 65.57 5.21 10.33 67.445.19 10.17 10  C₂₈H₂₈N₄O₉ 59.57 5.00 9.93 59.41 5.06 9.82 11  C₂₄H₂₄N₄O₇60.00 5.04 11.66 59.67 4.95 11.46

That which is claimed:
 1. A process for making a pharmaceutically activebis-aryl carbamate, said process comprising: reacting an aryl carbonatewith bis-amidine in the presence of an organic solvent to form thebis-aryl carbamate, wherein the pharmaceutically active bis-arylcarbamate may-se is represented by the formula:

wherein: X is O or S; R₁ and R₂ may be independently selected from thegroup consisting of H, loweralkyl, oxyalkyl, alkoxyalkyl, cycloalkyl,aryl, hydroxyalkyl, aminoalkyl, and alkylaminoalkyl; R₃ and R₄ are eachindependently selected from the group consisting of H, loweralkyl,halogen, oxyalkyl, oxyaryl, and oxyarylalkyl; R₅ is represented by aformula selected from the group consisting of:

wherein: X₁, X₂, and X₃ are independently selected from O and S; and R₆and R₇ are independently selected from the group consisting ofloweralkyl, aryl, alkylaryl, oxyaryl, an ester-containing substituent,and oxyalkyl.
 2. The process according to claim 1, wherein the arylcarbonate is selected from the group consisting of diphenyl carbonate,bis(4-fluorophenyl)carbonate, bis(4-methoxyphenyl)carbonate,benzyl-4-nitrophenylcarbonate, 4-nitrophenyl thioethyl carbonate, and4-nitrophenyl-2,2,2-trichloroethyl carbonate, methyl 4-nitrophenylcarbonate, bis (3-flourophenyl)carbonate, ethyl 4-nitrophenyl carbonate,(4-methyl-2-oxo-1,3-dioxol-4-en-5-yl)methyl 4-nitrophenyl carbonate, and1-acetoxyethyl 4-nitrophenyl carbonate.
 3. The process according toclaim 1, wherein the pharmaceutically active bis-aryl carbamate isselected from the group consisting of2,5-bis[4-(N-2,2,2-trichloroethoxycarbonyl)amidinophenyl]furan,2,5-bis[4-(N-thioethylcarbonyl)amidinophenyl]furan,2,5-bis[4-(N-benzyloxycarbonyl)amidinophenyl]furan,2,5-bis[4(N-phenoxycarbonyl)amidinophenyl]furan,2,5-bis[4-(N-(4-fluoro)phenoxycarbonyl)amidinophenyl]furan,2,5-bis[4-(N-(4-methoxy)phenoxycarbonyl)amidinophenyl]furan,2,5-bis[4(1-acetoxyethoxycarbonyl)amidinophenyl]furan, and2,5-bis[4-(N-(3-thio)phenoxycarbonyl) amidinophenyl]furan.
 4. Theprocess according to claim 1, wherein R₆ and R₇ are independentlyselected from the group consisting of: CH₃, CH₂CCl₃, CH₂CH₃,


5. The process according to claim 1, wherein each of the substituentspresent on the compound of formula (I) represented by the formula:

are present on the para positions of the aromatic groups on formula (I).6. The process according to claim 1, wherein the aryl carbonate isrepresented by the formula:

wherein: R is represented by:

wherein X is selected from the group consisting of H, NO₂, F, and OCH₁₃;and wherein R′ is selected from the group consisting of CH₃, CH₃CH₂,CH₂CCl₃, CH(OAc)CH₂, CH₂C₆H₅, and

wherein X is selected from the group consisting of H, NO₂, F, andOCH_(3.)
 7. The process according to claim 6, wherein the aryl carbonateis a symmetrical aryl carbonate.
 8. The process according to claim 1,wherein the organic solvent is selected from the group consisting ofdimethyl formamide and tetrahydrofuran/CH₃CN.
 9. The process accordingto claim 8, wherein the tetrahydrofuran/CH₃CN is employed in thepresence of a base.
 10. The process according to claim 9, wherein thebase is diisopropylethylamine.